title: False biomarker discovery due to reactivity of a commercial ELISA for CUZD1 with cancer antigen CA125.
abstract: BACKGROUND
  By using proteomics and bioinformatics, we have previously identified a group of highly pancreas-specific proteins as candidate pancreatic ductal adenocarcinoma (PDAC) biomarkers. With the use of commercially available ELISAs, the performance of some of these candidates was initially evaluated in a relatively small serum cohort (n = 100 samples). This phase revealed that CUB and zona pellucida-like domains protein 1 (CUZD1) may represent a new, promising PDAC biomarker.
  
  
  METHODS
  We performed detailed experiments to investigate the specificity of the commercial CUZD1 ELISA assay. CUZD1 was expressed in house in both bacteria and yeast expression systems. Recombinant CUZD1 and biological samples containing CUZD1, as well as commercial CUZD1 ELISA standards, were analyzed by Western blot, size exclusion HPLC, and mass spectrometry (LC-MS Orbitrap).
  
  
  RESULTS
  We confirmed that instead of CUZD1, the commercial assay is recognizing a nonhomologous, known cancer antigen [cancer antigen 125 (CA125)].
  
  
  CONCLUSIONS
  We conclude that poor characterization of commercial ELISA assays is a factor that could lead to false biomarker discovery. To our knowledge, this is the first report documenting that a commercial ELISA marketed for one analyte (CUZD1) may, in fact, recognize a different, nonhomologous antigen (CA125).
authors.affiliation: Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, ON, Canada;
authors.forename: Ioannis
authors.initials: I
authors.lastname: Prassas
authors.forename: Davor
authors.initials: D
authors.lastname: Brinc
authors.forename: Sofia
authors.initials: S
authors.lastname: Farkona
authors.forename: Felix
authors.initials: F
authors.lastname: Leung
authors.forename: Apostolos
authors.initials: A
authors.lastname: Dimitromanolakis
authors.forename: Caitlin C
authors.initials: CC
authors.lastname: Chrystoja
authors.forename: Randall
authors.initials: R
authors.lastname: Brand
authors.forename: Vathany
authors.initials: V
authors.lastname: Kulasingam
authors.forename: Ivan M
authors.initials: IM
authors.lastname: Blasutig
authors.forename: Eleftherios P
authors.initials: EP
authors.lastname: Diamandis
mesh_terms: D018394:CA-125 Antigen / Q000097:blood*; D002138:Calibration; D021441:Carcinoma, Pancreatic Ductal / Q000097:blood*; D045744:Cell Line, Tumor; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D055088:Early Detection of Cancer; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods / Q000592:standards*; D005188:False Negative Reactions; D006801:Humans; D008565:Membrane Proteins / Q000097:blood* / Q000235:genetics; D010190:Pancreatic Neoplasms / Q000097:blood*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D053719:Tandem Mass Spectrometry
chemical_list: D018394:CA-125 Antigen; C488236:CUZD1 protein, human; C451759:MUC16 protein, human; D008565:Membrane Proteins